12 直接门控传输:神经肌肉突触
我们对控制大脑化学突触的原理的大部分理解是基于对骨骼肌细胞上运动神经元形成的突触的研究。伯纳
· 卡茨及其同事从 1950 年开始的三十年间里程碑式的工作定义了突触传递的基本参数,并为现代突触功能的
分子分析打开了大门。因此,在我们研究中枢神经系统突触的复杂性之前,我们将研究更简单的神经肌肉突触
中化学突触传递的基本特征。
早期的研究利用了不同物种的神经肌肉制剂提供的几个实验优势。肌肉和附着的运动轴突很容易在体外解
剖并维持数小时。肌肉细胞足够大,可以被 2 或多个尖端微电极穿透,从而能够精确分析突触电位和潜在的
离子电流。在大多数物种中,神经元只在一个部位,即运动终板处受到支配,而在成年动物中,该部位仅受一个
运动轴突支配。相比之下,中枢神经元接收许多分布在整个树突末端和体细胞中的汇聚输入,因此单个输入的
影响更难辨别。
最重要的是, 20 世纪早期发现了介导神经和肌肉之间突触传递的化学递质乙酰胆碱我们现在知道,
经肌肉突触的信号传递涉及一个相对简单的机制:从突触前神经释放的神经递质与突触后膜中的一种受体结合,
即烟碱乙酰胆碱受体。1 递质与受体的结合直接打开一个离子通道;受体和通道都是同一大分子的组成部分。
活或抑制烟碱乙酰胆碱受体的合成和天然药物已被证明不仅可用于分析肌肉中的乙酰胆碱受体,而且可用于分
析外周神经节和大脑中的胆碱能突触。此外,此类配体可以是有用的治疗剂,包括治疗乙酰胆受体功能改
变或基因突变引起的遗传性和获得性神经系统疾病。
12.1 神经肌肉接头具有专门的突触前和突触后结构
当运动轴突接近终板时,即神经和肌肉之间的接触部位(也称为神经肌肉接头)它会失去髓鞘并分成几个
细小的分支。如图 12.1.1 所示,在它们的末端,这些细小的分支形成多次扩张或静脉曲张,称为突触神经节,
动轴突从中释放其递质。尽管髓磷脂的末端距离递质释放部位有一段距离,但施旺细胞覆盖并部分包裹神经末
梢。一个末端的“心轴”定义了运动终板的区域。在不同的物种中,终板的范围从约 20 微米的紧凑椭圆结构到
长度超过 100 微米的线性阵列。
神经末梢位于肌肉表面的沟槽中,即初级褶皱中。如图 12.1.1 所示,每个突触环下的膜进一步内陷形成一
系列次级或连接褶皱。神经末梢下方的肌肉细胞质包含许多可能参与突触特异性分子合成的圆形肌肉核。它们
不同于沿着肌纤维长度远离突触的扁平核。
轴突中的动作电位被传导到细分枝的末端,在那里它们触发乙酰胆碱释放。突触结节包含合成和释
酰胆碱所需的所有机制。这包括含有递质乙酰胆碱的突触小泡和突触小泡聚集的活动区。此外,每个活动区都
包含电压门控 Ca
2+
通道,这些通道通过每个动作电位将 Ca
2+
传导到末端。Ca
2+
的流入触发突触小泡与活性区
质膜的融合,通过胞吐作用将突触小泡的内容物释放到突触间隙中(第 15 章)
如图 12.1.4B 所示,可以使用 α-金环蛇毒素 (αBTX) 研究乙酰胆碱受体的分布,α-金环蛇毒素αBTX是一种
银环蛇的毒液中分离出来的肽,它与神经肌肉接头处的乙酰胆碱受体紧密且特异地结合。如图 12.1.2 所示,
BTX125I-αBTX的定量放射自显影显示乙酰胆碱受体堆积在次级褶皱的顶部,表面密度超过 10, 000m
2
负责定位受体的因素在第 48 章中讨论,我们将在其中考虑突触连接的发展。
神经肌肉接头处的突触前膜和突触后膜被 100 纳米宽的裂缝隔开。尽管拉蒙 · 卡哈尔 19 世纪末提出了
这样的差距,但直到 50 多年后用电子显微镜检查突触时才被可视化!整个突触间隙都存在由胶原蛋白和其他细
胞外基质蛋白组成的基底膜(基底层)酰胆碱酯酶是一种快速水解乙酰胆碱的酶,它被锚定在基底膜内的
胶原纤维上。释放到突触间隙中的乙酰胆碱在到达肌肉膜中的乙酰胆碱受体之前必须经过乙酰胆碱酯酶的“挑
战”由于乙酰胆碱酯酶被高浓度乙酰胆碱抑制,因此大多数分子都能通过。然而,该酶将乙酰胆碱的作用限制
为“一次击中”,因为乙酰胆碱酯酶一旦与突触后膜中的受体分离,就会水解递质。
12.1 神经肌肉接头具有专门的突触前和突触后结构
256 Part III / Synaptic Transmission
运动神经元
肌纤维
神经肌肉接头
髓磷脂
轴突
端板区域
施旺细胞鞘
突触扣结
线粒体
突触小泡 (
乙酰胆碱)
突触前膜
突触间隙
乙酰胆碱
接头褶皱
Figure 12–1 The neuromuscular junction is
an ideal site for studying chemical synaptic
signaling.At the muscle, the motor axon ram-
ifies into several fine branches approximately
2 μm thick. Each branch forms multiple
swellings called synaptic boutons, which are
covered by a thin layer of Schwann cells. The
boutons contact a specialized region of the
muscle fiber membrane, the end-plate, and
are separated from the muscle membrane by
a 100-nm synaptic cleft. Each bouton contains
mitochondria and synaptic vesicles clustered
around active zones, where the neurotransmit-
ter acetylcholine (ACh) is released. Immedi-
ately under each bouton in the end-plate are
several junctional folds, the crests of which
contain a high density of ACh receptors.
The muscle fiber and nerve terminal are
covered by a layer of connective tissue, the
basal lamina, consisting of collagen and glyco-
proteins. Unlike the cell membrane, the basal
lamina is freely permeable to ions and small
organic compounds, including the ACh trans-
mitter. Both the presynaptic terminal and the
muscle fiber secrete proteins into the basal
lamina, including the enzyme acetylcholinest-
erase, which inactivates the ACh released
from the presynaptic terminal by breaking
it down into acetate and choline. The basal
lamina also organizes the synapse by aligning
the presynaptic boutons with the postsynaptic
junctional folds.(Adapted from McMahan and
Kuffler 1971.)
Kandel-Ch12_0254-0272.indd 256 18/01/21 5:40 PM
受体通道
电压门控钠通道
突触后膜
活性区
钙离子通道
基板
12.1.1: 神经肌肉接头是研究化学突触信号的理想场所。在肌肉处,运动轴突分叉成几个约 2 微米厚的细分支。
每个分支形成多个肿块,称为突触环,上面覆盖着一层薄薄的施旺细胞结节接触肌纤维膜的一个特殊区域,
终板并通过 100 纳米的突触间隙与肌肉膜分开。每个结节都包含聚集在活跃区域周围的线粒体和突触小泡,
经递质酰胆碱这些区域被释放。紧接在终板每个环的下方是几个连接褶皱,其顶部含有高密度的乙酰胆碱
受体。肌肉纤维和神经末梢被一层结缔组织覆盖,基底层由胶原蛋白和糖蛋白组成。与细胞膜不同,基底层可自
由渗透离子和小的有机化合物,包括乙酰胆碱递质。突触前末端和肌纤维都将蛋白质分泌到基底层,包括乙酰
胆碱酯酶,它通过将突触前末端释放的乙酰胆碱分解成醋酸盐和胆碱来使乙酰胆碱失活。基底层还通过将突触
前环与突触后连接褶皱对齐来组织突触
[66]
219
12.1 神经肌肉接头具有专门的突触前和突触后结构
258 Part III / Synaptic Transmission
Figure 12–4 The end-plate potential can be isolated phar-
macologically for study.
A.Under normal circumstances, stimulation of the motor axon
produces an action potential in a skeletal muscle cell. The
dashed curve in the plot shows the inferred time course of the
end-plate potential that triggers the action potential. The lighter
dashed line shows the action potential threshold.
B.Curare blocks the binding of ACh to its receptor and so
prevents the end-plate potential from reaching the threshold
for an action potential. In this way, the currents and channels
that contribute to the end-plate potential, which are different
from those producing an action potential, can be studied.
The end-plate potential shown here was recorded in the
presence of a low concentration of curare, which blocks only
a fraction of the ACh receptors. The values for the resting
potential (–90 mV), end-plate potential, and action potential in
these intracellular recordings are typical of a vertebrate
skeletal muscle.
Figure 12–3 Acetylcholine receptors in
the vertebrate neuromuscular junction
are concentrated at the top one-third
of the junctional folds.This receptor-rich
region is characterized by an increased
density of the postjunctional membrane
(arrow). The autoradiograph shown here
was made by first incubating the membrane
with radiolabeled α-bungarotoxin, which
binds to the ACh receptor. Radioactive decay
results in the emittance of a particle that
causes overlaid silver grains to become fixed
along its trajectory (black grains). Magnifica-
tion ×18,000. (Reproduced, with permission,
from Salpeter 1987.)
By recording at different points along the muscle fiber,
Fatt and Katz found that the EPSP is maximal at the
end-plate and decreases progressively with distance
(Figure 12–5). In addition, the time course of the EPSP
slows progressively with distance.
From this, Fatt and Katz concluded that the end-
plate potential is generated by an inward ionic cur-
rent that is confined to the end-plate and then spreads
passively away. (An inward current corresponds to an
influx of positive charge, which depolarizes the inside
of the membrane.) Inward current is confined to the
end-plate because the ACh receptors are concentrated
there, opposite the presynaptic terminal from which
transmitter is released. The decrease in amplitude and
slowing of the EPSP as a function of distance is a result
of the passive cable properties of the muscle fiber.
Electrophysiological evidence that the ACh recep-
tors are localized to the end-plate was provided by
Stephen Kuffler and his colleagues, who applied
ACh to precise points on the muscle membrane
V
m
(mV)
–40
–60
Action potential
–20
0
+20
–80
+40
–90
A Normal
End-plate potential
Threshold
Time (ms)
5
01
0
V
m
(mV)
Time (ms)
–60
–70
–50
–80
–90
5
01
0
B With curare
Threshold
–40
–30
End-plate potential
Kandel-Ch12_0254-0272.indd 258 18/01/21 5:40 PM
12.1.2: 脊椎动物神经肌肉接头中的乙酰胆碱受体集中在连接褶皱的顶部 1/3 处。这个富含受体的区域的特
是连接后膜的密度增加(箭头)此处显示的放射自显影照片是通过首先将膜与放射性标记的 α-银环蛇毒素一起
孵育而制成的,它与乙酰胆受体结合。放射性衰变导致粒子发射,导致覆盖的银颗粒沿粒子轨迹叠加在一起
(黑色颗粒)。放大 18,000 倍。
12.1.1 膜通透性的局部变化导致突触后电位变化
一旦酰胆碱突触末稍释放,它会迅速结合并打开终板膜中乙酰胆受体通道。这导致肌肉膜对阳离
子的渗透性显著增加,导致正电荷进入肌肉纤维并使终板膜快速去极化。由此产生的兴奋性突触后电位非常大;
单个运动轴突的刺激产生大约 75 毫伏的兴奋性突触后电位在神经肌肉突触处,兴奋性突触后电位也称为终板
电位。
膜电位的这种变化通常大到足以快速激活肌肉膜中的电压门控 Na
+
通道,将终板电位转化为动作电位,然
后沿肌纤维传播。由于连接褶皱底部的电压门控 Na
+
通道密度很高,终板在肌肉中产生动作电位的阈值特别低。
一个非常大的奋性突触后电位低阈值的组合,使触发肌纤维动作电位的安全系数非常高。相反,在中枢神
经系统中,大多数突触前神经元产生的突触后电位幅度小于 1 毫伏,因此需要来自多个突触前神经元的输入才
能在大多数中枢神经元中产生动作电位。
保罗 · 法特伯纳德 · 卡茨 1950 年代首先使用细胞内电压记录详细研究了终板电位。法特卡茨能够通
过应用箭毒药物(图 12.1.4A来降低突触后电位的幅度,以其振幅低于动作电位的阈值,从而能够分离出终板
电位(图 12.1.5。在终板,突触电位在 1 2 毫秒内上升,但衰减得更慢。如图 12.1.3 所示,通过在肌肉纤维
的不同点进行记录,法特卡茨发现兴奋性突触后电位在终板处最大,并随着距离的增加而逐渐减小。此外,
奋性突触后电位的时间进程随着距离的增加而逐渐减慢。
由此,卡茨得出结论,终板电位是由局限在终板的内向离子电流产生的,然后这种电流被动地扩散
开来(内向电流对应于正电荷的流入,使膜内部去极化)内向电流被限制在终板,是因为乙酰胆碱受体集中在
那里,与释放递质的突触前末端相对应。作为距离的增加,兴奋性突触后电位振幅减小和速度减慢是肌肉纤维
的被动电缆特性的结果。
斯蒂芬 · 库夫勒及其同事他们使用一种称为微离子电渗疗法的技术将乙酰胆碱应用于肌肉膜上的精确点,
供了表明乙酰胆受体局限于终板的电生理学证据。在这种方法中,通过向电极内部施加正电压,带正电荷的
乙酰胆碱从充满乙酰胆碱的细胞外微电极中排出。将终板区域暴露于蛋白水解酶可使神经末梢从肌肉表面拉开,
并将乙酰胆碱直接应用于小微电极尖端正下方的突触后膜。使用这种技术,库夫勒发现在突触末稍几微米内,
220
12.1 神经肌肉接头具有专门的突触前和突触后结构
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 259
Figure 12–5 The end-plate potential
decreases with distance as it pas-
sively propagates away from the end-
plate.(Adapted, with permission, from Miles
1969.)
A.The amplitude of the postsynaptic potential
decreases and the time course of the potential
slows with distance from the site of initiation
in the end-plate.
B.The decay results from leakiness of the
muscle fiber membrane. Because charge must
flow in a complete circuit, the inward synaptic
current at the end-plate gives rise to a return
outward current through resting channels and
across the lipid bilayer (the capacitor). This
return outward flow of positive charge depolar-
izes the membrane. Because current leaks out
all along the membrane, the outward current
and resulting depolarization decreases with
distance from the end-plate.
A
B
0
123
4 毫米
运动
轴突
运动神经元终端
突触电流
肌纤维内部
肌纤维
using a technique called micro-iontophoresis. In this
approach, the positively charged ACh is ejected from
an ACh-filled extracellular microelectrode by apply-
ing a positive voltage to the inside of the electrode.
Exposing the end-plate region to proteolytic enzymes
allows the nerve terminal to be pulled away from the
muscle surface and the ACh to be applied directly to
the postsynaptic membrane directly under the tip of
the small microelectrode. Using this technique, Kuffler
found that the postsynaptic depolarizing response to
ACh declined steeply within a few micrometers of the
synaptic terminal.
Voltage-clamp experiments have revealed that the
end-plate current rises and decays more rapidly than
the resultant end-plate potential (Figure 12–6). The
time course of the end-plate current is directly deter-
mined by the rapid opening and closing of the ACh
receptor-channels. Because it takes time for an ionic
AB
Muscle
ber
+
Motor
axon
V
c
V
m
I
m
2 ms
Time
10 mV
10 nA
Stimulate motor axon
Depolarizing
end-plate potential
Inward synaptic current
at the end-plate
Figure 12–6 The end-plate current increases and decays
more rapidly than the end-plate potential.
A.The membrane at the end-plate is voltage-clamped by insert-
ing two microelectrodes into the muscle near the end-plate.
One electrode measures membrane potential (V
m
), and the
second passes current (I
m
). Both electrodes are connected to a
negative feedback amplifier, which ensures that sufficient cur-
rent (I
m
) is delivered so that V
m
will remain clamped at the com-
mand potential V
c
. The synaptic current evoked by stimulating
the motor nerve can then be measured at constant V
m
, for
example, −90 mV (see Box 10–1).
B.The end-plate potential (measured when V
m
is not clamped)
changes relatively slowly and lags behind the more rapid inward
synaptic current (measured under voltage-clamp conditions).
This is because synaptic current must first alter the charge on
the muscle membrane capacitance before the muscle
membrane can be depolarized.
Kandel-Ch12_0254-0272.indd 259 18/01/21 5:40 PM
12.1.3: 终板电势随着它被动的从终板处传播开去,振幅随距离减少。A. 突触后电位的振幅减小,并且电位的
时间进程随距离离开起始点而减慢。B. 这种衰减是由于肌肉纤维膜的泄漏导致的。因为电荷必须在一个完整的
回路中流动,所以终板处的内向突触电流会通过静息通道和脂质双层(即电容器)产生返回的外向电流。这种
返回的正电荷向外流动使膜去极化。因为电流沿着膜一直漏出,所以向外的电流和由此产生的去极化会随着与
终板的距离的增加而减小。
221
12.1 神经肌肉接头具有专门的突触前和突触后结构
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 257
Figure 12–2 Poisons, venoms, and high-voltage electric fish
help elucidate the structure and function of the nicotinic
ACh receptor.
A.Curare is a mixture of toxins extracted from the leaves of
Strychnos toxifera and is used by South American indigenous
people on arrowheads to paralyze their quarry. The active com-
pound, D-tubocurarine, is a complex multiring structure with
positively charged amine groups that bear some similarity to
ACh. It binds tightly to the ACh binding site on the nicotinic
receptor, where it acts as a competitive antagonist for ACh.
(Reproduced from Pabst, G (ed). 1898. Köhlers Medizinal-
Pflanzen, Vol. 3, Plate 45. Gera-Untermhaus, Germany: Franz
Eugen Köhler.)
B.The toxin α-bungarotoxin is obtained from the venom of
the banded krait, Bungarus. It is a 74-amino acid polypeptide
that contains five disulfide bonds (yellow lines), producing
a rigid structure (From https://en.wikipedia.org/wiki/Alpha-
bungarotoxin. Adapted from Zeng et al. 2001.). The toxin binds
extremely tightly to the ACh binding site and acts as an irrevers-
ible, noncompetitive antagonist of ACh.
C.The electric ray Torpedo marmorata has a specialized struc-
ture, the electric organ, which consists of a large number of
small, flat, muscle-like cells, or electroplaques, arranged in
series like a stack of batteries. When a motor nerve releases
ACh, a large current is generated by the opening of a very large
number of nicotinic ACh receptor-channels, which produces a
very large voltage drop of up to 200 V outside the fish, thereby
stunning nearby prey. The electroplaques provide a rich source
of ACh receptors for biochemical purification and characteriza-
tion.(From Walsh 1773.)
OCH
3
H
3
CCH
3
N
+
+
N
OCH
3
HCH
3
OH
OH
O
O
南美箭毒树
石纹电鳐
- 电鳐科
银环蛇
A C B
α-环蛇毒素
烟碱乙酰胆碱
D-
筒箭毒碱
channels in the muscle membrane, converting the end-
plate potential into an action potential, which then
propagates along the muscle fiber. The threshold for
generating an action potential in the muscle is particu-
larly low at the end-plate, owing to a high density of
voltage-gated Na
+
channels in the bottom of the junc-
tional folds. The combination of a very large EPSP and
low threshold results in a high safety factor for trigger-
ing an action potential in the muscle fiber. In contrast,
in the central nervous system, most presynaptic neu-
rons produce postsynaptic potentials less than 1 mV
in amplitude, such that inputs from many presynaptic
neurons are needed to generate an action potential in
most central neurons.
The end-plate potential was first studied in detail
in the 1950s by Paul Fatt and Bernard Katz using intra-
cellular voltage recordings. Fatt and Katz were able to
isolate the end-plate potential by applying the drug
curare (Figure 12–2A) to reduce the amplitude of the
postsynaptic potential below the threshold for the action
potential (Figure 12–4). At the end-plate, the synaptic
potential rises within 1 to 2 ms but decays more slowly.
Kandel-Ch12_0254-0272.indd 257 18/01/21 5:40 PM
12.1.4: 毒药、毒液和高压电鱼有助于阐明烟碱乙酰胆碱受体的结构和功能。A. 箭毒是从马钱子的叶子中提取
的毒素混合物,南美土著人涂在箭头上来麻痹他们的猎物。活性化合物 D-筒箭毒碱是一种复杂的多环结构化合
物,带有带正电荷的胺基,与乙酰胆碱有一些相似之处。它与烟碱受体上的乙酰胆碱结合位点紧密结合,作为
乙酰胆碱的竞争性拮抗剂发挥作用。B. α-环蛇毒素是从金环蛇(B ungarus)的毒液中提取的。它是一种包含
5 个二硫键(黄线)的 74 氨基酸多肽,产生了刚性结构。该毒素与乙酰胆碱结合位点极其紧密地结合,并作为
乙酰胆碱的不可逆、非竞争性拮抗剂发挥作用。C. 石纹电鳐有一个特殊的结构,即电器官,它由大量小而扁平
的肌肉状细胞或电斑组成,像电池堆一样串联排列。当运动神经释乙酰胆时,大量烟碱乙酰胆碱受体通道
的打开会产生大电流,从而在鱼体外产生高达 200 伏特的非常大的电压降,使附近的猎物昏迷。电板乙酰胆
受体的生化纯化和表征提供了丰富的来源。
222
12.1 神经肌肉接头具有专门的突触前和突触后结构
01
01
12.1.5: 终板电位可以通过药理学方法单独分离出来进行研究。A. 在正常情况下,运动轴突的刺激会在骨骼肌
细胞中产生一个动作电位。图中的虚线曲线显示了触发动作电位的终板电位的推断时间轨迹。较浅的虚线表示
动作电位的阈值。B. 箭毒阻断酰胆碱与其受体的结合,从而防止终板电位达到动作电位的阈值。通过这种方
式,有助于研究产生终板电位的电流和通道,这与产生动作电位的电流和通道不同。此处显示的终板电位是在
存在低浓度箭毒的情况下记录的,该浓度下箭毒仅阻断一部分乙酰胆碱受体。这些细胞内记录中的静息电位
90 毫伏)、终板电位和动作电位值是脊椎动物骨骼肌的特征。
乙酰胆碱的突触后去极化反应急剧下降。
如图 12.1.6 示,电压钳实验表明,终板电流的上升和衰减比由此产生的终板电势更快。终板电流的时间
过程直接乙酰胆受体通道的快速打开和关闭决定。因为离子电流对肌肉膜电容充电或放电需要时间,从而
改变膜电压,所以兴奋性突触后电位滞后于突触电流(参见图 9.6.1 和本章末尾的附言)
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 259
Figure 12–5 The end-plate potential
decreases with distance as it pas-
sively propagates away from the end-
plate.(Adapted, with permission, from Miles
1969.)
A.The amplitude of the postsynaptic potential
decreases and the time course of the potential
slows with distance from the site of initiation
in the end-plate.
B.The decay results from leakiness of the
muscle fiber membrane. Because charge must
flow in a complete circuit, the inward synaptic
current at the end-plate gives rise to a return
outward current through resting channels and
across the lipid bilayer (the capacitor). This
return outward flow of positive charge depolar-
izes the membrane. Because current leaks out
all along the membrane, the outward current
and resulting depolarization decreases with
distance from the end-plate.
A
B
0
1234 mm
Motor
axon
Motor neuron terminal
Synaptic current
Interior of muscle ber
Muscle
ber
using a technique called micro-iontophoresis. In this
approach, the positively charged ACh is ejected from
an ACh-filled extracellular microelectrode by apply-
ing a positive voltage to the inside of the electrode.
Exposing the end-plate region to proteolytic enzymes
allows the nerve terminal to be pulled away from the
muscle surface and the ACh to be applied directly to
the postsynaptic membrane directly under the tip of
the small microelectrode. Using this technique, Kuffler
found that the postsynaptic depolarizing response to
ACh declined steeply within a few micrometers of the
synaptic terminal.
Voltage-clamp experiments have revealed that the
end-plate current rises and decays more rapidly than
the resultant end-plate potential (Figure 12–6). The
time course of the end-plate current is directly deter-
mined by the rapid opening and closing of the ACh
receptor-channels. Because it takes time for an ionic
AB
肌纤维
+
运动
轴突
V
c
V
m
I
m
2 毫秒
时间
10 毫伏
10
钠安
刺激运动轴突
端板电位去极化
端板向内突触电流
Figure 12–6 The end-plate current increases and decays
more rapidly than the end-plate potential.
A.The membrane at the end-plate is voltage-clamped by insert-
ing two microelectrodes into the muscle near the end-plate.
One electrode measures membrane potential (V
m
), and the
second passes current (I
m
). Both electrodes are connected to a
negative feedback amplifier, which ensures that sufficient cur-
rent (I
m
) is delivered so that V
m
will remain clamped at the com-
mand potential V
c
. The synaptic current evoked by stimulating
the motor nerve can then be measured at constant V
m
, for
example, −90 mV (see Box 10–1).
B.The end-plate potential (measured when V
m
is not clamped)
changes relatively slowly and lags behind the more rapid inward
synaptic current (measured under voltage-clamp conditions).
This is because synaptic current must first alter the charge on
the muscle membrane capacitance before the muscle
membrane can be depolarized.
Kandel-Ch12_0254-0272.indd 259 18/01/21 5:40 PM
12.1.6: 终板电流比终板电势增加和衰减得更快。A. 通过将 2 个微电极插入终板附近的肌肉中,对终板处的膜
进行电压钳制。一个电极测量膜电位(V
m
,第二个电极测量电流(I
m
2 个电极都连接到负反馈放大器,以
确保输送足够的电流I
m
以便 V
m
保持钳位在命令电位 V
c
处。然后可以在恒定 V
m
下测量通过刺激运动神经
引发的突触电流,例如 −90 毫伏(见文本框 10.1B. 终板电位(在 V
m
未被钳制时测得)变化相对缓慢,滞后
于更快速的内向突触电流(在电压钳制条件下测得)这是因为在肌肉膜去极化之前,突触电流必须首先改变肌
肉膜电容上的电荷,然后肌肉膜才能去极化。
12.1.2 神经递质乙酰胆碱以离散包的形式释放
法特卡茨 1950 年代首次在青蛙运动终板进行微电极记录时,观察到小的自发去极化电位(0.51.0
伏),平均速率约为 1/s。这种自发电位仅限于终板区域,与刺激诱发兴奋性突触后电位表现出相同的时间过
程,并被箭毒阻断。因此,它们被命名为微终板电位
什么可以解释微型终板电位的小而固定的尺寸?德尔 · 斯蒂略卡茨测试了微兴奋性突触后电代表
乙酰胆分子作用的可能性。这个假设很快就被否定了,因为将非常少量的乙酰胆碱应用于终板可以引起远
小于 1.0-mV 微兴奋性突触后电位的去极化反应。低剂量的乙酰胆碱实会增加基准波动或“噪音”。后来对该
223
12.2 单个乙酰胆碱受体通道传导全有或无电流
噪声的统计成分的分析表明,潜在的单一突触后反应是振幅为 0.3 微伏且持续时间为 1.0 毫秒的去极化。这是单
乙酰胆碱受体通道(稍后描述)的电信号特性的第一个提示线索。
德尔 · 卡斯蒂略卡茨得出结论,每个微兴奋性突触后电位必须代表多分子包或“量子”递质的作用。此外,
他们认为由刺激诱发的大型兴奋性突触后电位由整数个量子组成。第 15 章介绍了该量子假说的证据。
12.2 单个乙酰胆碱受体通道传导全有或无电流
产生产生去极化终板电位的内向电流的乙酰胆碱受体通道的特性是什么?哪些离子通过通道移动以产生这
种内向电流?单个乙酰胆碱受体通道携带的电流是什么样的?
1976 年,厄温 · 内尔 · 克曼骨骼肌细胞中单乙酰胆受体通道传导的电流记录中获得了对
酰胆碱受体通道功能的生物物理性质的重要见解,即单一电流或基本电流。如图 12.2.1A 所示,他们发现,打开
单个通道会产生非常小的矩形离子电流脉冲。在给定的静息电位下,每个通道开口都会产生相同大小的电流脉
冲。 −90 毫伏时,电流阶跃的振幅约为 −2.7 皮安。虽然这是一个非常小的电流,但它相当于每秒约 1700 万个
离子的流动!
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 261
Figure 12–7 Individual acetylcholine (ACh) receptor-channels
conduct an all-or-none elementary current.
A.The patch-clamp technique is used to record currents from
single ACh receptor-channels. The patch electrode is filled with
salt solution that contains a low concentration of ACh and is
then brought into close contact with the surface of the mus-
cle membrane (see Box 8–1). At a fixed membrane potential,
each time a channel opens, it generates a relatively constant
elementary current. At the resting potential of –90 mV, the
current is approximately −2.7 pA (1 pA = 10
12
A). As the volt-
age across a patch of membrane is systematically varied, the
resultant current varies in amplitude as a result of changes in
driving force. The current is inward at voltages negative to 0 mV
and outward at voltages positive to 0 mV, thus defining 0 mV as
the reversal potential. The arrows on the right side of the traces
illustrate the individual sodium and potassium fluxes and
resultant net current as a function of voltage.
B.The linear relation between current through a single ACh
receptor-channel and membrane voltage shows that the chan-
nel behaves as a simple resistor having a single-channel con-
ductance (f) of about 30 pS.
膜电位
逆转电位
A B
+70 毫伏
–70 毫伏
100 毫秒
2 皮安
+55 毫伏
–50 毫伏
0 毫伏
60–40–8
08
04020–20–60
2.0
1.0
–1.0
–2.0
i
(皮安)
V
m
(
毫伏)
γ = 30 pS
乙酰胆碱受体通道
I
Na
+
I
K
净电流
size as the membrane potential changes because of the
change in driving force. For the ACh receptor-channels,
the relationship between I
EPSP
and membrane voltage is
linear, indicating that the single-channel conductance is
constant and does not depend on membrane voltage;
that is, the channel behaves as a simple ohmic resistor.
From the slope of this relation, the channel is found to
have a conductance of 30 pS (Figure 12–7B). As we saw in
Chapter 9, the total conductance, g, due to the opening of
a number of receptor-channels (n) is given by:
g = n × γ.
The current–voltage relation for a single chan-
nel shows that the reversal potential for ionic current
through ACh receptor-channels, obtained from the
intercept of the membrane voltage axis, is 0 mV, which
is not equal to the equilibrium potential for Na
+
or any
of the other major cations or anions. This is due to the
fact that this chemical potential is produced not by a
single ion species but by a combination of two species:
The ligand-gated channels at the end-plate are almost
equally permeable to both major cations, Na
+
and K
+
.
Thus, during the end-plate potential, Na
+
flows into
the cell and K
+
flows out. The reversal potential is at
0 mV because this is a weighted average of the equi-
librium potentials for Na
+
and K
+
(Box 12–1). At the
reversal potential, the influx of Na
+
is balanced by an
equal efflux of K
+
(Figure 12–7A).
The ACh receptor-channels at the end-plate are not
selective for a single ion species, as are the voltage-gated
Na
+
or K
+
channels, because the diameter of the pore of
the ACh receptor-channel is substantially larger than
that of the voltage-gated channels. Electrophysiological
measurements suggest that it may be 0.6 nm in diam-
eter, an estimate based on the size of the largest organic
cation that can permeate the channel. For example,
tetramethylammonium (TMA) is approximately 0.6 nm
in diameter and yet still permeates the channel. In con-
trast, the voltage-gated Na
+
channel is only permeant
to organic cations that are smaller than 0.5 × 0.3 nm in
cross section, and voltage-gated K
+
channels will only
conduct ions less than 0.3 nm in diameter.
The relatively large diameter of the ACh receptor-
channel pore is thought to provide a water-filled
Kandel-Ch12_0254-0272.indd 261 18/01/21 5:40 PM
12.2.1: 单个乙酰胆碱受体通道传导全有或全无的基本电流。A. 膜片钳技术用于记录单个乙酰胆碱受体通道的
电流。贴片电极充满含有低浓度乙酰胆的盐溶液,然后与肌肉膜表面紧密接触(见文本框 8.1。在固定的膜
电位下,每次通道打开时,它都会产生相对恒定的基本电流。90 毫伏的静息电位下,电流约为 −2.7 皮安(1
皮安 = 10
12
安)。随着膜片上的电压系统地变化,合成电流的幅度会因驱动力的变化而变化。电流在负电压至
0 毫伏时向内,在正电压至 0 毫伏时向外,因此将 0 毫伏定义为反转电位。迹线右侧的箭头说明了单独的钠和钾
通量以及作为电压函数的合成净电流。B. 通过单个乙酰胆碱受体通道的电流与膜电压之间的线性关系表明,该
通道表现为一个简单的电阻器,其单通道电导(f)约为 30 pS
12.2.1 终板的离子通道可渗透 Na
+
K
+
虽然通过单个酰胆碱体通道的电流幅度对于每次打开都是恒定的,但每次打开的持续时间和打开之间
的时间差异很大。这些变化是因为通道的打开和关闭,是随机的;它们遵循描述放射性衰变指数时间过程的相同
统计规律。由于通道乙酰胆会经历随机的热运动和波动,因此无法准确预测任何一个通道结乙酰胆
要多长时间,或者在乙酰胆解离和通道关闭之前该通道将保持打开状态多长时间。然而,特定类型通道保持
打开的平均时间长度是该通道的明确定义的属性,就像放射性衰变的半衰期是特定同位素的不变属性一样。
酰胆碱受体通道的平均开放时间约为 1 毫秒。因此,每个通道开口允许大约 17000 离子移动。一旦通道关闭,
乙酰胆碱分子就会解离,通道保持关闭状态,直到它再次与乙酰胆碱结合。
224
12.2 单个乙酰胆碱受体通道传导全有或无电流
一旦受体通道打开,哪些离子会流过通道,这如何导致肌肉膜去极化?识别负责突触电流的离子(或多个离
子)的一种重要方法是测量推动离子通过通道的化学驱动力(化学电池)的值。请记住,通过单个开放通道的电
流由单通道电导率与通过通道传导的离子的电化学驱动力的乘积给出(第 9 章)因此,单个乙酰胆碱受体通道
产生的电流由下式给出:
I
EPSP
= γ
EPSP
× (V
m
E
EPSP
), (12.1)
其中 I
EPSP
是通过一个通道的电流幅度,γ
EPSP
是单个开放通道的电导,E
EPSP
是膜电位,此时通过通道的离
子净通量 0V
m
E
EPSP
是离子通量的电化学驱动力。由于驱动力的变化,电流步长随着膜电位的变化而
化。对于乙酰胆碱受体通道,I
EPSP
与膜电压呈线性关系,表明单通道电导是恒定的,不依赖于膜电压;也就是
说,通道表现为一个简单的欧姆电阻。如图 12.2.1B 所示,根据该关系的斜率,发现通道的电导为 30 pS。正如
我们在第 9 章中看到的,由于许多受体通道(n)的开放,总电导 g 由下式给出:
g = n ×γ. (12.2)
单个通道的电流-压关系表明,通乙酰胆碱受体通道的离子电流的反转电位(从膜电压轴的截距获得)
0 毫伏,这不等于 Na
+
其他主要阳离子或阴离子任何的平衡电位。这是因为这种化学势不是由单一离子种类
产生,而是由 2 种离子种类的组合产生:终板上的配体门控通道几乎对 2 种主要阳离子 Na
+
K
+
具有同等的渗
透性。因此,在终板电位期间,Na
+
流入细胞,K
+
流出。反转电位为 0 毫伏,因为这是 Na
+
K
+
平衡电位的加
权平均值(文本框 12.1。如图 12.2.1A 所示,在反转电位处,Na
+
的流入被 K
+
的等量流出所平衡。
文本框 12.1 (终板电位的反转电位)
由一种以上离子物种携带的膜电流的反转电位,例如通过乙酰胆碱受体通道的终板电流,由 2 个因
素决定:1)渗透离子的相对电导(在终板电流的情况下为 g
Na
g
K
)和(2)离子的平衡电位(E
Na
E
K
在乙酰胆碱受体通道电流的反转电位下,Na
+
携带的内向电流与K
+
携带的外向电流平衡:
I
Na
+ I
K
= 0. (12.3)
单个Na
+
K
+
电流可以从以下 2 个方程获得:
I
Na
= g
Na
× (V
m
E
Na
) (12.4)
I
K
= g
K
× (V
m
E
K
). (12.5)
我们可以用等式 12.3 中的 I
Na
I
K
替换等式 12.4 12.5 E
EPSP
替换 V
m
(因为在反转电位 V
m
=
E
EPSP
时)
g
Na
× (E
EPSP
E
Na
) + g
K
× (E
EPSP
E
K
) = 0. (12.6)
求解 E
EPSP
的这个方程生成:
E
EPSP
=
g
Na
× E
Na
+ (g
K
× E
K
)
g
Na
+ g
K
. (12.7)
如果知道 E
EPSP
E
K
E
Na
,该方程也可用于求解 g
Na
/g
K
/比。因此,重新排列方程式 12.7 产生:
g
Na
g
K
=
E
EPSP
E
K
E
Na
E
EPSP
. (12.8)
在神经肌肉接头处,E
EPSP
=0 毫伏,E
K
=−100 毫伏,E
Na
=+55 毫伏。因此,从等式 12.8 可以看出,g
Na
/g
K
的值约为 1.8表明乙酰胆碱受体通道对Na
+
的电导略高于对K
+
的电导。一种可比较的方法可用于分析中
枢神经元兴奋性和抑制性突触电位期间的反转电位和离子运动(第 13 章)
终板上的乙酰胆碱受体通道对单一离子种类没有选择性,电压门控的 Na
+
K
+
通道也是如此,因为乙酰胆
受体通道的孔径远大于电压门控孔-门控通道。电生理测量表明它的直径可能 0.6 纳米,这是根据可以渗
225
12.3 乙酰胆碱受体通道具有与产生肌肉动作电位的电压门控通道不同的特性
透通道的最大有机阳离子的大小进行的估计的。例如,四甲胺的直径约为 0.6 纳米,但仍会渗透到通道中。相比
之下,电压门控 Na
+
通道只能透过横截面小于 0.5 × 0.3 纳米的有机阳离子,而电压门控 K
+
通道只能传导直径
小于 0.3 纳米的离子。
乙酰胆碱受体通道孔径的相对较大的原因被认为提供了一个充满水的环境,允许阳离子相对不受阻碍地通
过通道扩散,就像它们在游离溶液中一样。这解释了为什么孔不区分 Na
+
K
+
以及为什么即使是二价阳离子
(例如Ca
2+
)也能够通过。如本章后面所述,通道中存在固定负电荷,阴离子被排除在外。如图 12.2.2 所示,最
近的 X 射线晶体学数据提供了乙酰胆碱受体通道大孔的直接视图。
268 Part III / Synaptic Transmission
Figure 12–12 A high-resolution three-dimensional
structural model of a neuronal nicotinic ACh receptor-
channel.High-resolution models of the pentameric family of
ligand-gated channels are shown for the closed, open, and
desensitized states of the receptor-channel. Two out of five of
the M2 α-helixes are shown. The desensitized structure is from
the human neuronal ACh receptor. The closed and open states
are based on structures of neuronal glycine receptors, which
are closely related in amino acid sequence to ACh receptor sub-
units. Key amino acid side chains are illustrated for the desen-
sitized ACh receptor with position numbering on the right and
amino acid abbreviations on the left. According to convention,
position 0 is near the intracellular surface of the phospholipid
bilayer; other positions are labeled according to relative position
in the primary amino acid sequence. A conserved leucine in
the middle of the M2 segment (position 9) forms a gate that
constricts the pore in the closed state. Ligand binding causes
the subunits to tilt outward and twist, opening up the leucine
gate. A further conformational change during desensitization
causes the subunits to tilt inward near the bottom, constricting
the pore near the intracellular side of the channel and thereby
producing a nonconducting state. The negatively charged glu-
tamates at positions 20, –1, and –4 correspond to the external
(1), middle (2), and internal (3) rings of charge in Figure 12–11C.
The negatively charged glutamate at position –1 and the elec-
tronegative threonine at position 2 form the selectivity filter
of the channel. (Reproduced, with permission, from Morales-
Perez et al. 2016. Copyright © 2016 Springer Nature.)
2
–1
–4
20
9
Glu
Leu
LeuLeu
Thr
Glu
Glu
打开
脱敏
关闭
细胞外侧
细胞质侧
M3 transmembrane segments. This motion exerts a
force on the M2 segment that leads to its rotation and
tilting, thereby opening up the hydrophobic leucine
gate in the middle of the pore and allowing ion per-
meation. Although future studies will no doubt refine
our understanding of the structural bases for nicotinic
receptor-channel and function, these recent advances
give us an unprecedented molecular understanding
of one of the most fundamental processes in the nerv-
ous system: synaptic transmission and, specifically, the
signaling of information from nerve to muscle.
Highlights
1. The terminals of motor neurons form synapses
with muscle fibers at specialized regions in the
muscle membrane called end-plates. When an
action potential reaches the terminals of a presyn-
aptic motor neuron, it causes the release of ACh.
2. ACh diffuses across the narrow (100-nm) synaptic
cleft in a matter of microseconds and binds to nic-
otinic ACh receptors in the end-plate membrane.
The energy of binding is translated into a con-
formational change that opens a cation-selective
channel in the protein, allowing Na
+
, K
+
, and Ca
2+
to flow across the postsynaptic membrane. The
net effect, due largely to the influx of Na
+
ions,
produces a depolarizing synaptic potential called
the end-plate potential.
3. Because the ACh receptor-channels are concen-
trated at the end-plate, the opening of these chan-
nels produces a local depolarization. This local
depolarization is large enough (75 mV) to exceed
the threshold for action potential generation by a
factor of three to four.
4. It is important that the safety factor of nerve-
muscle transmission be at a high level, as it deter-
mines our ability to move, breath, and escape from
danger. Decreases in ACh receptor number or func-
tion as a result of autoimmune disease or genetic
mutations can contribute to neurological disorders.
5. Patch-clamp recordings have revealed the step-
like increase and decrease in current in response
to the opening and closing of single ACh receptor-
channels. A typical excitatory postsynaptic current
Kandel-Ch12_0254-0272.indd 268 18/01/21 5:40 PM
12.2.2: 神经元烟碱乙酰胆碱受体通道的高分辨率三维结构模型。显示了五聚体家族配体门控通道的高分辨率
模型,包括受体通道的关闭、打开和脱敏状态。显示了 5 M2 α-螺旋中的 2 个。脱敏结构来自人类神经元
酰胆碱体。关闭和打开状态基于神经元甘氨酸受体的结构,这些受体在氨基酸序列上乙酰胆受体亚基密
切相关。脱敏乙酰胆碱受体的关键氨基酸侧链在右侧显示,位置编号在左侧,氨基酸缩写在左侧。按照惯例,
0 位于磷脂双分子层的细胞内表面附近;其他位置根据一级氨基酸序列中的相对位置进行标记。M2 区段中间
(位置 9)的保守亮氨酸形成一个门,在关闭状态下收缩孔。配体结合导致亚基向外倾斜和扭曲,打开亮氨酸门。
脱敏过程中的进一步构象变化导致亚基在底部附近向内倾斜,收缩通道细胞内侧附近的孔,从而产生非导电状
态。位置 201 4 带负电荷的谷氨酸对应于图 12.3.3C 中的外部1中间2和内部3电荷环。位置
1 位带负电荷的谷氨酸和 2 位带负电的苏氨酸形成通道的选择性过滤器
[67]
12.2.2 四大因素决定终板电流
单个乙酰胆碱体通道携带的矩形电流阶跃如何在终板上产生大的突触电流以响应运动神经刺激?刺激运
动神经会释放大量乙酰胆碱进入突触间隙。乙酰胆碱迅速扩散穿过裂隙并与乙酰胆碱受体结合,导致超过 20
个受体通道几乎同时打开。(这个数字是通过比较总终板电流(大约 −500 纳安)与通过单个通道的电流(大约
−2.7 皮安)获得的)
这么多通道的快速打开导致终板膜的总电导 g
EPSP
大幅增加,并产生终板电流的快速上升阶段。由于间隙中
乙酰胆碱迅速减少至 0<1 毫秒)由于酶促水解和扩散,通道开始随机关闭。如图 12.2.3 所示,尽管每次闭
合仅使终板电流呈小的阶梯式下降,但随机闭合大量小的单一电流会导致总终板电流看起来平滑衰减。
12.3 乙酰胆碱受体通道具有与产生肌肉动作电位的电压门控通道不同的特性
产生终板电位的乙酰胆碱受体与产生肌肉动作电位的电压门控通道有 2 个重要区别。首先,动作电位是由 2
种不同类别的电压门控通道的顺序激活产生的,一种对 Na
+
具有选择性通透,另一种对 K
+
具有选择性通透。
比之下,单一类型的离子通道,乙酰胆碱受体通道,通过允许 Na
+
K
+
以几乎相等的渗透性通过而产生终板
电位。
226
12.3 乙酰胆碱受体通道具有与产生肌肉动作电位的电压门控通道不同的特性
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 263
muscle. First, the action potential is generated by
sequential activation of two distinct classes of voltage-
gated channels, one selective for Na
+
and the other for
K
+
. In contrast, a single type of ion channel, the ACh
receptor-channel, generates the end-plate potential by
allowing both Na
+
and K
+
to pass with nearly equal
permeability.
Second, the Na
+
flux through voltage-gated chan-
nels is regenerative: By increasing the depolarization
of the cell, the Na
+
influx opens more voltage-gated
A. Individual ACh receptor-channels open in response to a brief
pulse of ACh. In this idealized example, the membrane contains
six ACh receptor-channels, all of which open rapidly and nearly
simultaneously. The channels remain open for varying times
and close independently.
B. The stepped trace shows the sum of the six single-channel
current records in part A. It represents the current during the
sequential closing of each channel (the number indicates which
channel has closed). In the final period of current, only chan-
nel one is open. In a current record from a whole muscle fiber,
with thousands of channels, individual channel closings are not
detectable because the scale needed to display the total end-
plate current (hundreds of nanoamperes) is so large that the
contributions of individual channels cannot be resolved. As a
result, the total end-plate current appears to decay smoothly.
2
6
4
3
5
1
打开
关闭
通道 1
2
3
4
5
6
乙酰胆碱脉冲
2 毫秒
3 皮安
A 六个离子通道开放的理想时间过程
B 六个通道的总电流
2 毫秒
3 皮安
Kandel-Ch12_0254-0272.indd 263 18/01/21 5:40 PM
12.2.3: 终板总电流的时间进程反映了许多单独的乙酰胆碱受体通道贡献的总
[68]
A. 个体乙酰胆碱受体
道响应酰胆碱短暂脉冲而打开。在这个理想化的例子中,膜包含 6 乙酰胆碱受体通道,所有通道都快速
且几乎同时打开。这些通道在不同时间保持开放,并且独立关闭。B. 阶梯迹线为 A 部分 6 条单通道电流记录之
和,代表各通道依次关闭时的电流(数字表示关闭的通道)在电流的最后时期,只有一号通道是开放的。在整
个肌纤维的电流记录中,有数千个通道,无法检测到单个通道关闭,因为显示总终板电流(数百纳安)所需的比
例非常大,以至于无法解析出单个通道的贡献。因此,总终板电流看起来平稳衰减。
227
12.3 乙酰胆碱受体通道具有与产生肌肉动作电位的电压门控通道不同的特性
第二,通过电压门控通道 Na
+
流量是可再生的:通过增加细胞的去极化,Na
+
流入打开更多的电压门
Na
+
通道。这种再生特性决定了动作电位的全有或全无特性。相比之下,在突触电位期间打开的乙酰胆碱受体通
道的数量取决于可用酰胆碱数量。Na
+
通过酰胆碱门控通道流入产生的去极化不会导致更乙酰胆碱
体通道的打开,也不会产生动作电位。如图 12.3.1 所示,要触发动作电位,突触电位必须募集相邻的电压门控
Na
+
通道。
264 Part III / Synaptic Transmission
A minimal reaction model, first proposed by Katz
and his colleagues, captures many (but not all) of the
key steps of ACh receptor-channel function, in which a
closed receptor-channel (R) successively binds two mol-
ecules of ACh (A) prior to undergoing a rapid confor-
mational change to an open state (R*). This is followed
by a slower conformational change to the nonconduct-
ing desensitized state (D). The model also incorporates
the finding that there is a small probability that an indi-
vidual receptor may enter the desensitized state even in
the absence of ACh. These binding and gating reactions
can be summarized by the following scheme:
A + R AR + AA
2
RA
2
R*
A + D AD + AA
2
DA
2
D*
X-ray crystal structure models have now been
obtained for all three states of the ACh receptor
(described later).
The Low-Resolution Structure of the Acetylcholine
Receptor Is Revealed by Molecular and
Biophysical Studies
The nicotinic ACh receptor at the nerve-muscle syn-
apse is part of a single macromolecule that includes the
pore in the membrane through which ions flow. Where
in the molecule is the binding site located? How is the
Figure 12–9 The end-plate
potential resulting from the
opening of acetylcholine
receptor-channels opens
voltage-gated sodium chan-
nels.The end-plate potential
is normally large enough to
open a sufficient number of
voltage-gated Na
+
channels to
exceed the threshold for an
action potential.(Adapted from
Alberts et al. 1989.)
乙酰胆碱与
受体通道分子
的结合
Na
+
内流
K
+
外流
通道打开
去极化
(端板电位)
电压门控
Na
+
通道的打开
Na
+
内流
去极化
动作电位
电压门控
Na
+
通道
Na
+
乙酰胆碱
Na
+
乙酰胆碱
受体通道
K
+
pore of the channel formed? How is ACh binding cou-
pled to channel gating?
Insights into these questions have been obtained
from molecular and biophysical studies of the ACh
receptor proteins and their genes, beginning with the
purification of the macromolecule from the electric ray
Torpedo marmorata (Figure 12–2). Using different bio-
chemical approaches, Arthur Karlin and Jean Pierre
Changeux purified the receptor from electroplaques,
specialized muscle-like cells whose stack-like packing
enables their individual EPSPs to summate in series to
generate the large voltages (>100 V) used by the elec-
tric ray to stun its prey. Their studies indicate that the
mature nicotinic ACh receptor is a membrane glyco-
protein formed from five subunits of similar molecu-
lar weight: two α-subunits and one β-, one γ-, and one
δ-subunit (Figure 12–10).
Karlin and his colleagues identified two extracel-
lular binding sites for ACh on each receptor protein
in the clefts between each α-subunit and its neighbor-
ing γ- or δ-subunit. One molecule of ACh must bind at
each of the two sites for the channel to open efficiently
(Figure 12–10). Because α-bungarotoxin binds remark-
ably tightly to the same binding site on the α-subunit
as does ACh, the toxin acts as an irreversible transmit-
ter antagonist.
Further insights into the structure of the ACh
receptor-channel come from the analysis of the primary
amino acid sequence of the receptor’s four different
Kandel-Ch12_0254-0272.indd 264 18/01/21 5:40 PM
12.3.1: 乙酰胆碱受体通道开放导致的终板电位打开电压门控钠通道。终板电位通常足够大以打开足够数量的
电压门控 Na
+
通道以超过动作电位的阈值
[7]
正如从这 2 种生理特性的差异所预期的那样,乙酰胆碱受体通道和电压门控通道是由不同的大分子形成的,
这些大分子对药物和毒素表现出不同的敏感性。阻断电压门控 Na
+
通道的河豚毒素不会阻断 Na
+
通过烟碱乙酰
胆碱受体通道流入。同样,α-银环蛇毒素与烟碱受体紧密结合并阻断乙酰胆碱的作用,但不干扰电压门控 Na
+
K
+
通道。
12.3.1 递质结合在乙酰胆碱受体通道中产生一系列状态变化
每个乙酰胆碱受体都有 2 酰胆碱合位点;两者都必须被递质占用,通道才能有效打开。然而,在
酰胆碱的长期作用下,通道进入不再传导的脱敏状态。在正常情况下,肌肉烟碱受体脱敏的时间过程太慢,无
法影响兴奋性突触后电位的时间过程,其中酰胆碱存在于突触间隙中极短的时间内。然而,在某些神经元
突触中,脱敏可能在脱敏反应响应的过程中可以发挥更重要的作用。其中递质可能在突触间隙中持续更长时间,
或在突触后受体经历更快速的脱敏。
例如,乙酰胆碱在大脑胆碱能突触的突触间隙中的持续存在可能导致神经元烟碱受体的某些亚型显著脱敏。
重度吸烟者可以积聚足够量的尼古丁,使大脑中的受体脱敏。脱敏作用还在药物琥珀酰胆碱的作用中发挥作用,
琥珀酰胆碱是酰胆碱二聚体,对乙酰胆碱酯酶具有抗性,在全身麻醉期间用于产生肌肉松弛。琥珀胆碱通
过其产生受体脱敏和延长去极化的能力来实现这一点,这通过使电压门控 Na
+
通道失活来阻断肌肉动作电位。
228
12.3 乙酰胆碱受体通道具有与产生肌肉动作电位的电压门控通道不同的特性
卡茨其同事首先提出了最小反应模型,这个模型捕获了乙酰胆碱受体通道功能的许多(但不是全部)关
键步骤,其中一个闭合的受体通道(R,在经历快速构象变化到开放状态(R*)之前,依次结合 2 乙酰胆碱
分子A随后是受体通道的非传导脱敏状态D的较慢构象变化。该模型还考虑到以下发现,即使没有乙酰
胆碱,个体受体也有很小的可能进入脱敏状态。这些结合和门控反应可通过以下方案总结:
现在已经获得乙酰胆碱受体所有 3 种状态的 X 射线晶体结构模型(稍后描述)
12.3.2 分子和生物物理学研究揭示了乙酰胆碱受体的低分辨率结构
神经肌肉突触处的烟碱酰胆碱体是单个大分子的一部分,该大分子包括离子流经的膜孔。分子中的结
合位点位于何处?通道的孔道是如何形成的?乙酰胆碱结合如何与通道门控耦合?
如图 12.1.4 所示,乙酰胆受体蛋白及其基因的分子和生物物理学研究中获得了对这些问题的见解,首
先是从电射线石纹电鳐中纯化大分子。通过使用不同的生化方法,亚瑟 · 卡林 · 皮埃尔 · 尚热从电斑中纯化
了受体,电斑是专门的肌肉样细胞,其堆叠状包装使它们各自的奋性突触后电位联求和以产生大电压(大
100 伏特),用于电击猎物。如图 12.3.2 所示,他们的研究表明,成熟的烟乙酰胆受体是一种膜糖蛋白,
5 个分子量相似的亚基组成:2 α-亚基和 1 β-亚基、1 γ-亚基和 1 δ-亚基。
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 265
subunits and from biophysical studies. Molecular
cloning by Shosaku Numa and colleagues demon-
strated that the four subunits are encoded by dis-
tinct but related genes. Sequence comparison of the
subunits shows a high degree of similarity—one-half
of the amino acid residues are identical or conserva-
tively substituted—which suggests that all subunits
have a similar structure. Furthermore, all four of the
genes for the subunits are homologous; that is, they
are derived from a common ancestral gene. Nicotinic
ACh receptors in neurons are encoded by a set of
distinct but related genes. All of these receptors are
pentamers; however, their subunit composition and
stoichiometry vary. Whereas most neuronal receptors
are composed of two α-subunits and three β-subunits,
some neuronal receptors are composed of five identi-
cal α -subunits (the α7 isoform) and so can bind five
molecules of ACh.
All nicotinic ACh receptor subunits contain a
highly conserved sequence near the extracellular bind-
ing site for ACh consisting of two disulfide-bonded
cysteine (cys) residues with 13 intervening amino
acids. The resultant 15-amino acid loop forms a sig-
nature sequence both for nicotinic ACh receptor sub-
units and for related receptors for other transmitters
in neurons. The cys-loop receptor family, also known as
pentameric ligand-gated ion channels (pLGIC), includes
Figure 12–10 The nicotinic ACh receptor-channel is a
pentameric macromolecule.The receptor and channel are
components of a single macromolecule consisting of five
subunits: two identical α-subunits and one each of β-, γ-, and
δ-subunits. The subunits form a pore through the cell mem-
brane. When two molecules of ACh bind to the extracellular
binding sites—formed at the interfaces of the two α-subunits
and their neighboring γ- and δ-subunits—the conformation of
the receptor-channel molecule changes (see Figure 12–12). This
change opens the pore through which K
+
and Na
+
flow down
their electrochemical gradients.
Na
+
K
+
β
α
α
γ
δ
γ
乙酰胆碱
没有乙酰胆碱结合:通道关闭 两个乙酰胆碱分子结合:
通道打开
α
γ
α
α
αα
乙酰胆碱结合位点
receptors for the neurotransmitters γ-aminobutyric
acid (GABA), glycine, and serotonin.
The distribution of the polar and nonpolar amino
acids of the subunits provided the first clues as to
how the subunits are threaded through the mem-
brane bilayer. Each subunit contains four hydrophobic
regions of approximately 20 amino acids called M1
to M4, each of which forms an α-helix that spans the
membrane (Figure 12–11A). The amino acid sequences
of the subunits suggest that the subunits are arranged
such that they create a central pore through the mem-
brane (Figure 12–11B).
The walls of the channel pore are formed by the
M2 membrane-spanning segment and by the loop con-
necting M2 to M3. Three rings of negative charges that
flank the external and internal boundaries of the M2
segment play an important role in the channel’s selec-
tivity for cations. Certain local anesthetic drugs block
the channel by interacting with one ring of polar ser-
ine residues and two rings of hydrophobic residues in
the central region of the M2 helix, midway through the
membrane.
Three-dimensional models of the entire receptor-
channel complex were initially proposed by Karlin
based on low-resolution neutron scattering and by
Nigel Unwin based on electron diffraction images.
The complex is divided into three regions: a large
Kandel-Ch12_0254-0272.indd 265 18/01/21 5:40 PM
12.3.2: 烟碱型乙酰胆碱受体通道是一种五聚体大分子。受体和通道是单个大分子的组成,该大分子由 5 个亚基
组成:2 个相同的 α-亚基和 β-亚基、γ-亚基和 δ -亚基各一个。这些亚基形成一个穿过细胞膜的孔道。如图 12.2.2
示,当 2 乙酰胆碱分子与细胞外结合位点结合时(在 2 α-亚基及其相邻的 γ-亚基和 δ-亚基的界面处形成)
受体通道分子的构象发生变化。这种变化打开了 K
+
Na
+
沿其电化学梯度向下流动的孔隙,使 K
+
Na
+
顺着
其电化学梯度流动。
卡林和他的同事在每个 α-亚基与其相邻的 γ-亚基或 δ-亚基之间的裂隙中的每个受体蛋白上确定了乙酰胆碱
2 个细胞外结合位点。如图 12.3.2 所示,2 个位点里,每个位点都必须结合一个乙酰胆碱分子,通道才能有效
打开。由于 α -银环蛇毒素与乙酰胆碱一样与 α-亚基上的相同结合位点紧密结合,因此该毒素作为不可逆的递质
拮抗剂。
乙酰胆受体通道结构的进一步了解来自对受体 4 不同亚基的一级氨基酸序列的分析和生物物理学研
究。沼正作及其同事的分子克隆实验表明, 4 个亚基由不同但相关的基因编码。亚基的序列比较显示出高度
的相似性(一半的氨基酸残基相同或被保守取代)这表明所有亚基都具有相似的结构。此外,所有 4 个亚基基
因都是同源的;也就是说,它们源自共同的祖先基因。神经元中的烟乙酰胆受体由一组不同但相关的基因
编码。所有这些受体都是五聚体;然而,它们的亚基组成和化学计量比各不相同。虽然大多数神经元受体由 2
229
12.3 乙酰胆碱受体通道具有与产生肌肉动作电位的电压门控通道不同的特性
α 亚基和 3 β 亚基组成,而一些神经元受体由 5 个相同的 α 亚基(α7 亚型)组成,因此可以结合 5 乙酰胆
分子。
所有烟碱乙酰胆碱受体亚基在乙酰胆碱的细胞外结合位点附近都包含一个高度保守的序列,该序列由 2
二硫键结合半胱氨酸残基和他们之间 13 个氨基酸组成。由此产生的 15 氨基酸环形成了烟碱酰胆碱受体
基和神经元中其他递质的相关受体的特征序列。半胱氨环受体家族,也称五聚体配体门控离子通道包括
神经递质 γ-氨基丁酸、甘氨酸和 5-羟色氨的受体。
亚基的极性和非极性氨基酸的分布为我们提供了关于亚基如何穿过膜双层的第一条线索。如 12.3.3A
示,每个亚基包含 4 个大约由 20 个氨基酸组成的疏水区域,称为 M1 M4每个区域形成一个跨膜的 α 螺旋。
如图 12.3.3B 所示,亚基的氨基酸序列表明,亚基的排列使得它们在膜上形成一个中心孔。
266 Part III / Synaptic Transmission
Figure 12–11 The ACh receptor subunits are homologous
membrane-spanning proteins.
A.Each subunit contains a large extracellular N-terminus, four
membrane-spanning α-helixes (M1–M4), and a short extracel-
lular C-terminus. The N-terminus contains the ACh-binding site,
and the membrane helixes form the pore.
B.The five subunits are arranged such that they form a central
aqueous channel, with the M2 segment of each subunit form-
ing the lining of the pore. The γ-subunit lies between the two
α-subunits. (Dimensions are not to scale.)
C.Negatively charged amino acids on each subunit form three
rings of negative charge around the pore. As an ion traverses
the channel, it encounters these rings of charge. The rings
at the external and internal surfaces of the cell membrane
(1, 3) may serve as prefilters that help repel anions and form
divalent cation blocking sites. The central ring near the cyto-
plasmic side of the membrane bilayer (2) may contribute more
importantly to establishing the specific cation selectivity of
the selectivity filter, which is the narrowest region of the pore.
D.A high-resolution X-ray crystal structure model of a
human neuronal nicotinic ACh receptor-channel. Right:
A top-down view of the open channel, which is composed
of two
α
4
-subunits and three β
2
-subunits arranged around
the central pore. These subunits are closely related
variants of the α- and β-subunits of the muscle receptor.
Two molecules of nicotine (atoms shown as red spheres)
are bound to the receptor. A permeating cation is shown as
a pink sphere.Center: A side view of the receptor showing
the location of the phospholipid bilayer of the membrane
and bound nicotine. Left: A side view of a single α
4
-subunit
in the plane of the membrane. The amino-terminus of the
subunit consists of a large extracellular domain. Loop C
helps form the ligand-binding site. The β
1
-β
2
and cys-loops
at the interface between the extracellular domain and the
M1–M4 membrane-spanning α-helixes transmit a confor-
mational change from the ligand-binding site to the pore
to open the channel.(Reproduced, with permission, from
Morales-Perez et al. 2016. Copyright © 2016 Springer
Nature.)
A 乙酰胆碱受体通道中的单个亚
D
NH
2
COOH
细胞外侧
细胞质侧
细胞外侧
细胞质侧
δ
α
αγ
β
M1
M2
M4 M3
M3M1 M4M2
C 乙酰胆碱受体通道的功能模型
细胞外侧
漏斗形
入口区域
出口区 离子选择性孔隙
烟碱
细胞质侧
δ
αβ
––
––
1
2
3
B 通道孔周围亚基的排列
β2
β2
β1-β2 loop
M1
M4
M2
M2-M3 loop
Loop C
Cys-loop
M3
C
β2
α4
α4
N
α4
Kandel-Ch12_0254-0272.indd 266 18/01/21 5:40 PM
12.3.3: 乙酰胆受体亚基是同源的跨膜蛋白。A. 每个亚基都包含一个大的细胞外 N 末端、4 跨膜 α 螺旋
M1M4和一个短的细胞外 C 末端。N 端包含乙酰胆碱结合位点,膜螺旋形成孔。B. 5 个亚基的排列使得它们
形成一个中央水通道,每个亚基的 M2 部分形成孔的内层。γ 亚基位于 2 α 亚基之间(尺寸未按比例绘制)C.
每个亚基上带负电荷的氨基酸在孔周围形 3 个带负电荷的环。当离子穿过通道时,它会遇到这些电荷环。细
胞膜内外表面的环1, 3可用作预滤器,帮助排斥阴离子并形成二价阳离子阻断位点。膜双层2细胞质侧附
近的中心环可能更重要地有助于建立选择性过滤器的特定阳离子选择性,这是孔的最窄区域。D. 人类神经元烟
乙酰胆碱受体通道的高分辨率 X 射线晶体结构模型。右图:开放通道的俯视图,由围绕中心孔排列的 2 a4
亚基和 3 b2 亚基组成。这些亚基是肌肉受体的 α β 亚基的密切相关变体。2 个尼古丁分子(显示为红色球
体的原子)与受体结合。一个渗透阳离子显示为粉红色球体。中心:受体的侧视图,显示膜的磷脂双层和结合尼
古丁的位置。左图:膜平面中单个 α 亚基的侧视图。该亚基的氨基末端由一个大的细胞外结构域组成。环 C
助于形成配体结合位点。胞外域和 M1-M4 跨膜 α 螺旋之间界面处的 β1-β2 和半胱氨酸环将构象变化从配体
合位点传递到孔以打开通道
[67]
230
12.4 亮点
通道孔壁由 M2 跨膜段和连接 M2 M3 的环路形成。位于 M2 区段内外边界两侧的 3 个负电荷环在通道对
阳离子的选择性中起着重要作用。某些局部麻醉药通过与膜中途 M2 螺旋中心区域的一个极性丝氨酸残基环和
2 个疏水性残基环相互作用来阻断通道。
整个受体通道复合体的三维模型最初由卡林基于低分辨率中子散射和奈杰尔 · 昂温基于电子衍射图像提出。
如图 12.3.3C 所示,该复合体分为 3 个区域:包含乙酰胆碱结合位点的大细胞外部分、对阳离子具有选择性的窄
跨膜孔和内膜表面的大出口区域。细胞外区域非常大,长度约为 6 纳米。此外,孔的细胞外端有一个直径约为
2.5 纳米的宽口。在膜的双层内,孔逐渐变窄。
自身免疫性疾病重症肌无力是由与乙酰胆碱受体胞外结构域结合的抗体的产生引起的,导致神经肌肉接头
处烟碱型乙酰胆碱受体的数量或功能下降。如果变化足够严重,这可能会将奋性突触后电位低到触发动作
电位的阈值以下,从而导致虚弱无力。几种先天性肌无力形式是由烟碱乙酰胆碱受体亚基突变引起的,这些亚
基突变也可以改变受体数量或通道功能。例如,M2 区段氨基酸残基的突变会导致通道开放时间延长,称为慢通
道综合症,这会导致过度的突触后兴奋,从而导致终板退化(第 57 章)
12.3.3 乙酰胆碱受体通道的高分辨率结构通过 X 射线晶体研究揭示
胆碱位点节的深入解最自软动物酰胆结合的高辨率 X 线晶学研
究,该蛋白与烟碱乙酰胆碱受体亚基的细胞外氨基末端同源。值得注意的是,与典型的乙酰胆碱受体不同,软体
动物乙酰胆碱结合蛋白是一种由神经胶质细胞分泌到细胞外空间的可溶性蛋白。在蜗牛的胆碱能突触中,它可
以减小兴奋性突触后电位的大小,这可能是通过缓冲突触间隙中乙酰胆碱的游离浓度来减小的。
对完受体结构进一了解自于和多胞动的相五聚配体通道 X 射线
构。最近的一项研究揭示了与尼古丁复合的人神经元烟碱乙酰胆碱受体的 X 射线晶体结构。结合相关蛋白质的
结构知识,我们现在乙酰胆受体通道和相关配体门控通道的配体结合、通道门控和离子渗透的结构和机制
有了非常详细的了解。
如图 12.3.3D 所示,在神经元乙酰胆碱受体中,2 α-亚基与 3 β-亚基结合形成五聚体。受体的大细胞外
结构域包含 2 乙酰胆碱结合位点,并形成一个五聚体环,围绕着一个大的中央前庭,这可能将离子汇集到受体
的狭窄跨膜结构域。每个 α-亚基在位于与相邻 β-亚基界面的位点结合一个尼古丁分子。如图 12.2.2 所示,来自
奈杰尔 · 的相半胱氨酸受体的高分辨率结构和神经元烟碱受体脱敏状态的高分辨率结构的电子衍射
据表明,每个亚基的四个跨膜区段确实是以 α 螺旋的形式穿过 3 纳米长的脂质双层的长度。在脱敏状态下,来
5 个亚基的 M2 区段在膜的细胞内侧附近形成狭窄的收缩,阻止离子渗透。
我们对处于打开和关闭状态的烟碱乙酰胆碱受体通道的跨膜区域的理解仍然不完整。然而,通过与相
聚体配体门控离子通道的结构相比,受体的一致图像开始出现。在闭合状态下,孔道里的 M2 段大致彼此平行,
形成一个狭窄的中央孔。如图 12.2.2 所示,M2 区段中间附近高度的保守的疏水性亮氨酸残基将孔进一步收缩至
直径 0.3 0.4 纳米。这种疏水收缩被认为提供了一个高能屏障,限制直径大于孔道收缩的水合阳离子的通过。
目前,从电生理测量(0.6 纳米)推断的孔道直径与从晶体结构得出的较窄值之间的差异仍未解决。
在开放状态下,M2 片段被认为向外倾斜并旋转,扩大了 M2 中间亮氨酸残基的收缩,从而允许离子渗透。
开放孔中最窄的收缩部分位于通道的细胞内口附近,其中来自一个苏氨酸残基环(肌肉乙酰胆碱受体中的丝氨
酸和苏氨酸残基)的电负性羟基侧链和第二个带负电荷的谷氨酸残基环被认为形成选择性滤波器。在脱敏状态
下,
M2
部分进一步倾斜,导致选择性过滤器进一步收缩,从而阻止离子渗透。
基于各种结构和功能研究,现在正在逐渐形成配体结合如何导致通道开放的详细图景。配体的结合被认为
可以促进相邻亚基之间裂缝的闭合,从而导致五聚体的细胞外结构域收紧,类似于花瓣的闭合。这导致一个扭
转运动,导致受体细胞外结构域的底部推动 M1 区段和连接 M2 M3 跨膜区段的细胞外环。该运动对 M2 片段
施加一个力,导致其旋转和倾斜,从而打开孔中间的疏水亮氨酸门并允许离子渗透。尽管未来的研究无疑将进
一步完善我们对烟碱受体通道和功能的结构基础的理解,但这些最新进展使我们对神经系统中最基本的过程之
一有了前所未有的分子理解:突触传递,特别是信号传导从神经到肌肉的信息传递。
231
12.4 亮点
12.4 亮点
1. 在称为终板的肌膜特殊区域,运动神经元的末端与肌纤维形成突触。当动作电位到达突触前运动神经元
的末端时,它会导致乙酰胆碱的释放。
2. 乙酰胆碱散穿过狭窄的(100 纳米)突触间隙,只需要几微秒,并与终板膜中的烟碱酰胆碱受体结
合。结合能量转化为构象变化,打开蛋白质中的阳离子选择性通道,允许 Na
+
K
+
Ca
2+
流过突触后膜。主要
由于 Na
+
的流入,产生了一种去极化突触电位,称为终板电位。
3. 因为乙酰胆碱受体通道集中在终板,这些通道的打开会产生局部去极化。这种局部去极化足够大(75
伏),超过了动作电位产生的阈值 3 4 倍。
4. 神经肌肉传递的安全系数必须保持在很高的水平,因为它决定了我们移动、呼吸和逃生的能力。自身免
疫性疾病或基因突变导致的乙酰胆碱受体数量或功能下降可能导致神经系统疾病。
5. 膜片钳记录显示,在单个乙酰胆碱受体通道的打开和关闭时,电流呈阶梯式增加和减少。神经肌肉接
处,一个典型的兴奋性突触后电流是由大约 20 万个单独通道的开放产生的。
6. 肌肉烟碱乙酰胆碱受体的生化结构已经确定。该受体是由 2 α-亚基和 1 β-γ- δ-亚基组成的五聚体。
编码这些亚基的四个基因密切相关,与编码其他递质的其他五聚体配体门控通道的基因关系较远。
7. 更高分辨率的结构提供乙酰胆碱配体结合口袋和通道孔的详细视图,并进一步揭示配体结合如何导致
与受体通道开放和脱敏门控反应相关的构象变化。
12.5 后记:终板电流可从等效回路中计算
通过许多个乙酰胆碱受体通道的电流可以用欧姆定律来描述。然而,为了描述电流如何产生终板电位,还
必须考虑周围膜中静息通道的电导。我们还必须考虑膜的电容特性和由细胞内外的 Na
+
K
+
分布决定的离
电势差。
这些不同组件之间的动态关系可以使用相同规则来解释。我们在 9 章中用于分析仅由细胞膜及其通道的
电学特性和电势差组成的电流通路中的电流。我们可以用一个具有 3 个并联电流通路的等效回路来表示终板区
域:1一个用于通过递质门控通道的突触电流,2一个用于通过静息通道(非突触膜)的返回电流,3
一个用于跨越脂质双层的电容电流(图 12.5.1。为简单起见,我们忽略了周围非突触膜中的电压门控通道。
因为终板电流是通过同一离子通道的 Na
+
K
+
流动而传输的,我们将 Na
+
K
+
电流通路合并为一个单一
的电导g
EPSP
代表乙酰胆碱受体通道。该通路的电导率与打开的通道数成正比,而打开的通道数又取决于突
触间隙中递质的浓度。在没有递质的情况下,没有通道打开并且电导为 0当一个递质释放的前突触动作电位导
乙酰胆碱释放时,该通路的电导增加到大约 5 ×10
6
S大约是代表静息(泄漏)通道(g
l
的并联分支电导
5 倍。
如图 12.5.1 所示,终板电导与细胞E
EPSP
串联,其值由突触电流0 毫伏)的反转电位给出。该值是 Na
+
K
+
平衡电位的加权代数和(见文本框 12.1。兴奋性突触后电位(I
EPSP
)期间的电流由下式给出
L
EPSP
= g
EPSP
× (V
m
E
EPSP
). (12.9)
使用这个等式和图 12.5.1 的等效回路,我们现在可以根据其组件分析兴奋性突触后电位(图 12.5.2
兴奋性突触后电位始时(动态阶段),因为对 Na
+
K
+
的电导增加以及在 −90 毫伏静息电位下对 Na
+
的大内向驱动力,一种内向电流I
EPSP
流过乙酰胆碱受体通道(图 12.5.2B时间 2由于电荷在闭环中流动,
内向突触电流通过两条并行的通路离开细胞,成为通过静息(或泄漏)通道的外向电流:一条是离子电流(I
l
通过静息(或泄漏)通道,另一条是电容电流(I
c
)穿过脂质双分子层。
因此,
I
EPSP
= (I
l
+ I
c
). (12.10)
232
12.5 后记:终板电流可从等效回路中计算
Chapter 12 / Directly Gated Transmission: The Nerve-Muscle Synapse 269
at the neuromuscular junction is generated by
the opening of approximately 200,000 individual
channels.
6. The biochemical structure of the muscle nicotinic
ACh receptor has been determined. The receptor is
a pentamer composed of two α-subunits and one
β-γ-, and δ-subunit. The four genes encoding the
subunits are closely related, and more distantly
related to the genes encoding other pentameric
ligand-gated channels for other transmitters.
7. Higher-resolution structures have provided a
detailed view of the ACh ligand-binding pocket
and the pore of the channel and further insight
into how ligand binding leads to conformational
changes associated with receptor-channel opening
and desensitization gating reactions.
Postscript: The End-Plate Current Can Be
Calculated From an Equivalent Circuit
The current through a population of ACh receptor-
channels can be described by Ohm’s law. However,
to describe how the current generates the end-plate
potential, the conductance of the resting channels in
the surrounding membrane must also be considered.
We must also take into consideration the capacitive
properties of the membrane and the ionic batteries
determined by the distribution of Na
+
and K
+
inside
and outside the cell.
The dynamic relationships between these various
components can be explained using the same rules we
used in Chapter 9 to analyze the current in passive elec-
trical devices that consist only of resistors, capacitors,
and batteries. We can represent the end-plate region
with an equivalent circuit that has three parallel cur-
rent paths: (1) one for the synaptic current through the
transmitter-gated channels, (2) one for the return cur-
rent through resting channels (the nonsynaptic mem-
brane), and (3) one for the capacitive current across the
lipid bilayer (Figure 12–13). For simplicity, we ignore
the voltage-gated channels in the surrounding nonsyn-
aptic membrane.
Because the end-plate current is carried by both
Na
+
and K
+
flowing through the same ion channel,
we combine the Na
+
and K
+
current pathways into
a single conductance (g
EPSP
) representing the ACh
receptor-channels. The conductance of this pathway
is proportional to the number of channels opened,
which in turn depends on the concentration of trans-
mitter in the synaptic cleft. In the absence of transmit-
ter, no channels are open and the conductance is zero.
When a presynaptic action potential causes the release
of ACh, the conductance of this pathway increases to
approximately 5 × 10
−6
S, which is about five times the
conductance of the parallel branch representing the
resting (leakage) channels (g
l
).
The end-plate conductance is in series with a bat-
tery (E
EPSP
) with a value given by the reversal potential
for synaptic current (0 mV) (Figure 12–13). This value
is the weighted algebraic sum of the Na
+
and K
+
equi-
librium potentials (see Box 12–1). The current during
the excitatory postsynaptic potential (I
EPSP
) is given by
I
EPSP
= g
EPSP
× (V
m
E
EPSP
).
Using this equation and the equivalent circuit of
Figure 12–13, we can now analyze the EPSP in terms of
its components (Figure 12–14).
At the onset of the EPSP (the dynamic phase),
an inward current (I
EPSP
) flows through the ACh
C
m
g
EPSP
=
5.0 × 10
–6
S
g
l
=
1.0 × 10
–6
S
+
+
E
EPSP
(0 毫伏)
E
l
(–90 毫伏)
V
m
=
–15 毫伏
细胞外侧
细胞质侧
突触膜中的通道
非突触膜
中的通道
I
开关
+
+
V
Figure 12–13 The equivalent circuit of the end-plate.The
circuit has three parallel current pathways. One conductance
pathway carries the end-plate current and consists of a battery
(E
EPSP
) in series with the conductance of the ACh receptor-
channels (g
EPSP
). Another conductance pathway carries current
through the nonsynaptic membrane and consists of a battery
representing the resting potential (E
l
) in series with the con-
ductance of the resting channels (g
l
). In parallel with both of
these conductance pathways is the membrane capacitance
(C
m
). The voltmeter (V) measures the potential difference
between the inside and the outside of the cell.
When no ACh is present, the ACh receptor-channels are
closed and carry no current. This state is depicted as an open
electrical circuit in which the synaptic conductance is not con-
nected to the rest of the circuit. The binding of ACh opens the
synaptic channels. This event is electrically equivalent to throw-
ing the switch that connects the gated conductance pathway
(g
EPSP
) with the resting pathway (g
l
). In the steady state, an
inward current through the ACh receptor-channels is balanced
by an outward current through the resting channels. With the
indicated values of conductances and batteries, the membrane
will depolarize from −90 mV (its resting potential) to −15 mV
(the peak of the end-plate potential).
Kandel-Ch12_0254-0272.indd 269 18/01/21 5:40 PM
12.5.1: 终板的等效回路。该回路具有 3 个平行的电流。一种电导通路承载终板电流,由串联的乙酰胆碱受体
在突触后膜上产生的电势差与乙酰胆碱受体g
EPSP
的电导(E
EPSP
组成。另一个电导路径通过非突触膜传输
电流,由代表静息电位E
l
的细胞静息电势与静息通道的电导g
l
串联组成。与这两条电导通路并联的是膜
电容C
m
电压表V测量细胞内外之间的电位差。当不存在乙酰胆碱时,乙酰胆碱受体通道关闭并且不传
导电流。这种状态被描述为一个开式的回路,在这个回路中,突触电导没有连接到回路的其余部分。乙酰胆碱
结合打开突触通道。此事件在电气上等效于打开连接门控电导通路g
EPSP
与静息通路g
l
的开关。在稳定状
态下,通过乙酰胆受体通道的内向电流与通过静息通道的外向电流平衡。根据所示的电导和电池值,膜将从
−90 毫伏(其静息电位)去极化至 −15 毫伏(终板电位的峰值)
兴奋性突触后电位
的最早阶段,膜电位
V
m
仍接近其静息值
E
l
因此,电流通过静止通道
V
m
E
l
向外驱动力很小。因此,大部分外向电流以电容电流的形式离开细胞,膜迅速去极化(图 12.5.2B时间 2
着细胞去极化,通过静息通道的电流向外驱动力增加,而通过乙酰胆碱受体通道的突触电流向内驱动力减小。
时,随着突触中酰胆碱度的降低,乙酰胆碱受体通道开始关闭,最终通过门控通道的内向电流与通过静息
通道的外向电流完全平衡(I
EPSP
= I
l
。此时,没有电荷流入或流出电容器(I
c
= 0。因为膜电位的变化率与
I
c
成正比,
I
c
/C
m
= V
m
/t, (12.11)
膜电位将达到峰值或新的稳态值,V
m
/t = 0(图 12.5.2B,时间 3
随着乙酰胆碱受体通道关闭,I
EPSP
进一步降低。现在 I
EPSP
I
l
不再处于平衡状态,膜电位开始复极化,
为通过泄漏通道I
l
的外向电流变得大于内向突触电流。在突触动作电位作用的下降阶段的大部分时间里,
酰胆碱受体通道都没有电流,因为它们都已经关闭了。相反,电流仅通过膜传导作为由静息通道携带的外向电
流,并由内向电容电流平衡(图 12.5.2B,时间 4
兴奋性突触后电位处于峰值或稳态值时,I
c
= 0,因此可以轻松计算出 V
m
的值。通过乙酰胆碱受体通道
I
EPSP
)的内向电流必须与通过静息通道(I
l
)的外向电流完全平衡:
I
EPSP
+ I
1
= 0. (12.12)
通过乙酰胆碱受体通道(I
EPSP
)和静息通道(I
l
)的电流由欧姆定律给出:
I
EPSP
= g
EPSP
× (V
m
E
EPSP
), (12.13)
I
l
= g
l
× (V
m
E
l
). (12.14)
233
12.5 后记:终板电流可从等效回路中计算
270 Part III / Synaptic Transmission
Figure 12–14 The time course of the end-plate potential
is determined by both the ACh-gated synaptic conductance
and the passive membrane properties of the
muscle cell.
A.The time course of the end-plate potential and the compo-
nent currents through the ACh receptor-channels (I
EPSP
), the
resting (or leakage) channels (I
l
), and the capacitor (I
c
). There
is a capacitive current only when the membrane potential is
changing. In the steady state, such as at the peak of the end-
plate potential, the inward flow of positive charge through the
ACh receptor-channels is exactly balanced by the outward ionic
current across the resting channels, and there is no capacitive
current.
B.Equivalent circuits for the current at times 1, 2, 3, and
4 shown in part A. (The relative magnitude of a current is
represented by the arrow length.)
C
m
I
EPSP
= 0
E
EPSP
(0
毫伏)
细胞质侧
突触膜中的通道
非突触膜中的通道
E
l
(–90
毫伏)
g
l
g
EPSP
时间
1
V
c
=
Q
C
E
EPSP
E
l
I
EPSP
I
l
I
c
I
c
I
c
= 0I
EPSP
= –I
l
I
l
E
EPSP
E
l
时间 3
稳态
(突触作用的峰值)
I
l
I
EPSP
E
EPSP
E
l
I
l
时间 4
动态
(突触作用减弱)
I
EPSP
= 0
1
2
3
4
2 毫秒
10 毫伏
I
EPSP
I
c
A
B
+
+
++
––
++
––
+
+
+
+
+
+
+
+
+
++
––
+
终板电位
电流
稳态
(静息电位)
I
l
= 0 I
c
= 0
细胞外侧
时间 2
动态
(突触作用的开始)
I
EPSP
= I
l
+ I
c
Kandel-Ch12_0254-0272.indd 270 18/01/21 5:40 PM
12.5.2: 终板电位的时间进程由乙酰胆碱门控突触电导和肌肉细胞的被动膜特性决定。A. 终板电位和通过乙酰
胆碱受体通道(I
EPSP
、静息(或泄漏)通道(I
l
)和电容器(I
c
)的分量电流的时间过程。只有当膜电位发生
变化时才会有电容电流。在稳态下,例如在终板电位的峰值处,通过乙酰胆碱受体通道的内向正电荷流与通过
静息通道的外向离子电流完全平衡,并且没有电容电流。B. A 分所示时间 123 4 电流的等效回路(电
流的相对大小由箭头长度表示)
234
12.5 后记:终板电流可从等效回路中计算
将这 2 个表达式代入公式 12.12,我们得到:
g
EPSP
× (V
m
E
EPSP
) + g
l
× (V
m
E
l
) = 0. (12.15)
求解 V
m
,我们得到:
V
m
=
(g
EPSP
× E
EPSP
) + (g
l
× E
l
)
g
EPSP
+ g
l
. (12.16)
这个方程类似于用于计算静息电位和动作电位的方程(第 9 章)根据公式 12.16兴奋性突触后电位的峰
值电压是乙酰胆碱体通道和静息(泄漏)通道的 2 细胞的电动势的加权平均值。加权因子由 2 个电导的
对大小给出。由于 g
l
是一个常数,g
EPSP
的值越大(即乙酰胆碱通道开放的越多)V
m
将越接近 E
EPSP
的值。
我们现在可以计算图 12.5.1 中所示特定情况的峰值兴奋性突触后电位其中 g
EPSP
= 5 ×10
6
Sg
l
= 1 ×10
6
SE
EPSP
= 0 毫伏,E
l
= −90 毫伏。将这些值代入公式 12.16 可得:
V
m
=
[(5 × 10
6
S) × (0 mV)] + [(1 × 10
6
S) × (90 mV)]
(5 × 10
6
S) + (1 × 10
6
S)
(12.17)
或者
V
m
=
(1 × 10
6
S) × (90 mV)
(6 × 10
6
S)
= 15 mV. (12.18)
兴奋性突触后电位的峰值振幅为:
δ = V
m
E
1
= 15 mV (90 mV) = 75. (12.19)
235